The longterm objective of this research program is to determine the neuroendocrine mechanisms regulating LH and prolactin (PRL) secretion in mammals. Proposed studies examine hypothalamic mechanisms by which hyperprolactinemia (HP) and testosterone (T) may inhibit tonic LE secretion in male rats. This problem is of central importance in the control of mammalian fertility and has profound contraceptive significance. We will utilize an acute, reversible hyperprolactinemia model where there is a dose-related, transitory suppression of the postcastration rise in LH levels produced by ovine PRL (oPRL) administration. This animal model allows us to investigate the direct brain and pituitary effects of hyperprolactinemia on tonic LH release in the absence of confounding gonadal influences. We will analyze the effects of selected, increasing doses of administered oPRL on the frequency and amplitude of pulsatile LH secretion. Norepinephrine (NE), epinephrine, dopamine and GABA synthesis and turnover rate constants will be determined in intact, castrated, castrated-PRL-treated and castrated-T-treated rats, in microdissected brain regions which are likely sites of synapse of these neurons on LH-RH neurons. Quantitative in situ hybridization histochemistry will be utilized to determine whether PRL and T affect single cell levels of glutamic acid decarboxylase mRNA in medial preoptic area (MPoA) GABAergic neurons. Whether HP inhibits median eminence LH-RH turnover estimated by a new colchicine method, or blocks the LH response to third ventricular NE, MPOA infused bicuculline or iv N-methyl-D-aspartate will be ascertained. In this reversible model, brain parameters will be determined before (Oh),' during (48h) and after (96h) PRL suppression of the postcastration LH rise, thus assessing which endpoints change at a time of maximal suppression (48h) and whether these changes reverse or others appear at a time of reversal (96h). This research will further our understanding of the hypothalamic mechanisms regulating LH and PRL secretion and the ways in which elevated circulating PRL levels and feedback steroid affect these mechanisms.